Dear Ragoo, Several replies already mentioned possible causes for the change of appearance. For this same reason we use a milder and more physiological activation for lymphocytes using antibodies against CD2 (or CD3 ) and CD28. Some references: 1. R.A.W. van Lier et al.: 'Functional studies with anti-CD3 heavy chain isotype switch-variant monoclonal antibodies. Accessory cell-independent induction of lnterleukin-2 responsiveness in T cells by epsilon anti-CD3. J.Immunol. 139:2873 (1987). 2. R.A.W. van Lier et al.: 'Signals involved in T cell activation. T cell proliferation induced through the synergistic action of anti-CD28 and anti-CD2 monoclonal antibodies'. Eur.J.Immunol. 18:167 (1988). 3. H.M. Kuiper et al.: 'Influence of CD28 costimulation on cytokine production is mainly regulated via IL-2'. Immunology 83:38 (1994). The antibodies are available at Sanquin, www.clb.nl <http://www.clb.nl> Best regards, John John Voorn Sanquin Reagents Product manager Plesmanlaan 125 j_voorn@clb.nl <mailto:j_voorn@clb.nl> 1066CX AMSTERDAM the Netherlands tel +31-(0)20 5123246 fax +31-(0)20 5123570 web site www.clb.nl <www.clb.nl> -----Original Message----- From: Arumugham Raghunathan [SMTP:ragoor@molecularstaging.com] Sent: Monday, May 06, 2002 10:56 PM To: Cytometry Mailing List Subject: Lymphocyte activation Hi all, A question about activation of lymphs. When I activated lymphocytes using PMA and ionomycin, I see a population of cells with reduced side scatter properties apart from the regular lymphocytes. It looks like there are two populations (parallel shift) based on SSC. The shift is towards the left, and the number of cells shifting increases with activation-time. Has anyone observed this? What is it due to? Do the cells (lymphs) shrink upon activation? thanks, Ragoo *********************************************************** Arumugham Raghunathan, Ph.D Flow Cytometry Group Cellular Analysis Section Molecular Staging Inc., 300 George Street, Suite 701 New Haven, CT 06510 Tel# 203-772-5074 Fax# 203-776-5278 *************************************************************
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